Urea → 2-Hydroxyethyl Methacrylate

Participants

Reaction Steps

Aspergillus oryzae strain SE01-28 described in Example 9 was transformed with pSE38 to create new transformants designated Aspergillus oryzae SE27 according to the method described in Example 8 with the exceptions that Basta resistance was used for selection and 2-8 μg of NdeI digested pSE38 was added per transformation directly from the reaction so that ~5 U of enzyme was included in the transformation mixture. Media for Basta selection contained 20 ml of COVE salts, 1 M sucrose, 25 g/L Noble agar, 10 mM Urea, and either 5 mg/ml Basta (Hoechst Schering, Rodovre, Denmark) for transformation or 10 mg/ml Basta for maintaining transformants. The COVE salts were composed of 26 g of KCl, 26 g of MgSO4 -7H2O, 76 g of KH2PO4, and 50 ml of COVE trace elements per liter of deionized water. The COVE trace elements were composed of 0.04 g of Na2B4O7 -10H2O, 0.4 g of CuSO4 -5H2O, 1.2 g of FeSO4 -7H2O, 0.7 g of MnSO4 -H2O, 0.8 g of Na2MoO2 -H2O, 10 g of ZnSO4 -7H2O per liter of deionized water. Maltose was added to a final concentration of 2.5%, when indicated. Basta selection overlay medium was the same as above except that Basta was added prior to use.

Provenance & Evidence